Although adenovirus based gene therapy is capable of highly efficient gene transfer into nondividing cells, particular t1, respiratory epithelium, acute inflammation and acquired immune responses to the virus and to virus-infected cells have become a significant limitation. Our preliminary studies suggest that production of proinflammatory cytokines in general, and tumor necrosis factor alpha (TNF alpha) in particular, play a key role in the early inflammatory cell influx, and subsequent tissue injury and apoptotic cell death in the lung. This application proposes that modifying the early innate immune response to adenovirus gene transfer can 1) reduce the inflammatory cell infiltration and parenchymal injury that occurs after adenovirus transfection in lung, and 2) alter the cytotoxic T-cell and antibody reactions which limit transgene duration. In particular, we propose that coexpression of either inhibitors of the proinflammatory cytokine response or their downstream signal pathways can reduce the inflammatory properties of adenovirus, block apoptosis of virally-infected cells, and potentially extend the duration of transgene expression. The initial goal of this application is to examine first in mice and subsequently in primate (Papio) whether coexpression of either the anti-inflammatory cytokine IL-10, a pox virus derived soluble TNF receptor homologue, myxoma derived serine protease inhibitor (serp1), or a rabbitpox derived promiscuous chemokine (C-C, C-X-C) receptor, p35, in first- generated adenovirus constructs, can reduce the proinflammatory cytokine, lung innury, antibody and CTL responses, and extend duration of transgene expression when administered intratracheally. Preliminary data also suggests that death of virally-infected epithelial cells involves TNF alpha depednet apoptotic cell processes. Therefore, the second overally goal is to determine whether coexpression of the cowpox-virus derived gene product, CrmA, a member of the serpin family with concomitant cysteine-protease inhibitor, can reduce cysteine proteinase mediated apoptotic cell death in mice intransasally administered first generation adenovirus CrmA constructs. Inhibition of caspase-1 activity by CrmA overexpression should also reduce the apoptotic death of virus infected cells and extend transgene expression.